1. Safety in Chemical Laboratories

2. Introduction 1. A chemical lab is potentially hazardous environment 2. Accident and injury can happen anytime 3. Lab safety is everyone’s responsibility 4. Lab safety standards and practices must be strictly followed

5. Fire Extinguisher

6. DNA Extraction and Purification

7. Lab Equipments (To be used in this experiment) Automatic pipettes Microcentrifuge Vortex Water bath UV-spectrophotometer

8. Steps in DNA Extraction and Purification

11. Quantification of the purified DNA

12. Spectrophotometer Most of visible spectrophotometers are composed of: ◦ Light source which works with visible wavelengths (400-700 nm) ◦ Monochromator filter for choosing desired wavelength ◦ Sample holder (cuvette) ◦ Detector ◦ Meter or recorder

13. Measurements ◦ Dilute the sample of purified DNA: Add 700 μL of AE to the purified DNA ◦ Measure the Absorbance at 260nm ◦ Measure the Absorbance at 280nm ◦ Assess the DNA purity: 260/280 ratio (Accepted ratio: 1.7 - 1.9) ◦ Calculate DNA Conc.: Provided A260 = 1.0, DNA is 50 μg /ml, unknown DNA conc. can be calculated by cross multiplication A260 = 1.0 DNA conc. = 50 μg /ml A260 = 0.5 DNA conc. ?  Calculate DNA Yield: DNA Volume x DNA conc. Done by the spectrophotometer

14. DNA Yield: ◦ If you have Volume of DNA solution: 100 μ l (0.1 ml) DNA conc.: 30 μg /ml Then the yield ( μg ) = Volume x Conc. 0.1 ml x 30 μg /ml = 3.0 μ g

15. DNA Applications Purified DNA can be used for: 1. Molecular diagnosis of diseases (e.g., sickle cell anemia) 2. Forensic applications (e.g., paternity testing) 3. Molecular biology research

16. Molecular techniques using purified DNA: a. Amplification techniques: polymerase chain reaction (PCR) b. Southern blotting c. Restriction Fragment length polymorphism (RFLP)